Solid phase peptide synthesis was introduced in 1963 with the intent to overcome many of the intermediate purification problems associated with solution peptide synthesis. Stewart, et al., Solid Phase Peptide Synthesis (Pierce Chemical Co., 2d ed., 1984). During solid phase synthesis, amino acids are assembled (i.e., coupled) into a peptide of any desired sequence while one end of the chain (i.e., the C-terminus) is anchored to an insoluble support. Once the desired sequence has been linked together on the support, the peptide is then deblocked (i.e., cleaved) from the support. The two standard protecting groups for α-amino functions of the coupled amino acids are Boc, which is removed by treatment with a strong acid, and Fmoc, which is removed with a base. The present invention relates to a convenient method of manufacturing peptides using a combination of both of these α-amino protecting groups in a single synthesis on inexpensive chloromethylated polystyrene resin.
In designing a synthesis of a peptide by the solid phase method using either of the above mentioned α-amino protection schemes, it is important that any reactive “side groups” of the constituent amino acids be protected from unwanted chemical reactions throughout the chain assembly. It is also desirable that the chemical groups chosen to protect the various side groups be resistant to removal by the reagents used to remove the α-amino protecting groups. Thirdly, it is important that the linkage of the growing peptide chain to the resin particle be stable to the reagents used to remove either type of α-amino protecting group during chain assembly. In the case of the Fmoc α-amino protection scheme, the side group protection functions should be resistant to the basic reagents used to remove the Fmoc. In practice, these side chain protecting groups are generally removed by mildly acidic reagents after the peptide chain has been assembled. When using the Boc α-amino protection scheme, the side chain protecting groups must be resistant to removal by the mild acid reagent used to deprotect the Boc group at every cycle. In practice, these side chain protecting groups for the Boc α-amino protection scheme are generally removed by anhydrous HF after the peptide chain has been assembled. Therefore, in practice, the side chain protecting groups commonly used with the Fmoc α-amino protection are not stable under the conditions used for Boc α-amino deprotection and the two types of α-amino protection schemes are not mixed in the assembly of a peptide chain by solid phase peptide synthesis. In addition, while the least expensive polymeric resin used in peptide synthesis, chloromethylated polystyrene or “Merrifield resin”, is widely used with Boc protected amino acids, the literature suggests it is unsuitable for use with Fmoc protection on the α-amino group due to its lability in basic conditions. (see Stewart, et al., Solid Phase Peptide Synthesis (Pierce Chemical Co., 2d ed., 1984). The present invention is directed to a method for mixed usage of both Boc and Fmoc amino acids on “Merrifield resin” during solid phase synthesis of certain peptides.
Lanreotide® is an analog of somatostatin and is known to inhibit growth hormone release as well as inhibit insulin, glucagon and pancreatic exocrine secretion.
U.S. Pat. No. 4,853,371 discloses and claims Lanreotide®, a method for making it and a method for inhibiting the secretion of growth hormone, insulin, glucagon and pancreatic exocrine secretion.
U.S. Pat. No. 5,147,856 discloses the use of Lanreotide® of treating restenosis.
U.S. Pat. No. 5,411,943 discloses the use of Lanreotide® for treating hepatoma.
U.S. Pat. No. 5,073,541 discloses the use of Lanreotide® for treating lung cancer.
U.S. application Ser. No. 08/089,410 filed Jul. 9, 1993 discloses the use of Lanreotide® for treating melanoma.
U.S. Pat. No. 5,504,069 discloses the use of Lanreotide® for inhibiting the accelerated growth of a solid tumor.
U.S. application Ser. No. 08/854,941 filed May 13, 1997, discloses the use of Lanreotide® for decreasing body weight.
U.S. application Ser. No. 08/854,943 filed May 13, 1997, discloses the use of Lanreotide® for treating insulin resistance and Syndrome X.
U.S. Pat. No. 5,688,418 discloses the use of Lanreotide® for prolonging the survival of pancreatic cells.
PCT Application No. PCT/US97/14154 discloses the use of Lanreotide® for treating fibrosis.
U.S. application Ser. No. 08/855,311 filed May 13, 1997, discloses the use of Lanreotide® for treating hyperlipidemia.
U.S. application Ser. No. 08/440,061 filed May 12, 1995, discloses the use of Lanreotide® for treating hyperamylinemia.
U.S. application Ser. No. 08/852,221 filed May 7, 1997, discloses the use of Lanreotide® for treating hyperprolactinemia and prolactinomas.
The contents of the foregoing patents and patent applications are incorporated herein by reference.